Fig 6. Rational design and computational analysis of PY motif peptide library to predict residue-specific changes in binding affinity.

(A) Nedd4 WW domain (PDB ID: 2KPZ) in complex with PY motif peptides from previously resolved WW domain/PY peptide complexes (PDB IDs: 2KPZ, 2M3O, 2KQ0, 4N7H). Peptides aligned to the 2KPZ complex using PyMol [75], showing moderate conservation of peptide backbone conformation when bound to the WW domain. (B,C) Rational design of PY peptide library involved variation of residues in the x_–1 and x positions (shown in pink, B) and was informed by PY motif consensus sequences for Nedd4 (shown in Fig 2C and S2 Fig), affording a 30 member library. (D) Computationally predicted binding affinities of PY motif peptides screened against Nedd4 WW domain (PDB ID: 2M3O). Binding affinities are presented as ΔΔG_binding relative to the native peptide substrate TAPPPAYATLG (ΔG_binding^designed − ΔG_binding^native). ΔΔG_binding for the native peptide is presented in the upper-right corner of the left heatmap for reference. ΔΔG_binding energies presented as kcal/mol or % of ΔG_binding^native. Full energy properties provided as in S1 Data.